Effect of Glucose on Endo-xylanase and β-xylosidase Production by Fungi Isolated in Indonesia

Xylanases are widely produced by fungi, and the production of polysaccharide-degrading enzymes, in general, are usually subjected to carbon catabolite repression. In this work, the ability of several Indonesian indigenous fungi to produce endo-xylanase and β-xylosidase and their responses to glucose as a repressor were determined. Ten fungi were grown in a liquid medium supplemented with glucose as the repressor (0, 1%, 3%, and 5%), and the endo-xylanase and β-xylosidase productions were assayed. Aspergillus aculeatus FIG1 and A. oryzae KKB4 produced 3.85 and 0.70 U/mL of endo-xylanase, respectively, compared with other strains (0.22 U/mL or less). Trichoderma asperellum PK1J2, T. virens MLT2J2, A. aculeatus FIG1, T. asperellum MLT5J1, A. oryzae KKB4, and T. asperellum MLT3J2 produced 0.021–0.065 U/mL of β-xylosidase, whereas the other strains produced 0.013 U/mL or less of β-xylosidase. Adding 1% glucose to the growth medium can partially repress endo-xylanase production in A. aculeatus FIG1, T. asperellum PK1J2, and T. virens MLT4J1 and completely repress other strains. By adding 1% glucose, strains FIG1, PK1J2, and MLT4J1 suffered almost complete repression of β-xylosidase production, although such strains exhibited partial repression of endo-xylanase production. β-Xylosidase produced by the other strains showed complete repression by adding 1% glucose, except for A. aculeatus FIG1, A. tamarii FNCC 6151, and T. asperellum MLT1J1, which showed partial repression. Therefore, adding 3% glucose to the growth medium can result in complete repression of endo-xylanase and β-xylosidase productions in all strains examined.

Transcriptomic Analysis of the White-rot Basidiomycete Lentinus Squarrosulus to Provide Insights Into Its Lignocellulose Biodegradation Ability

BackgroundBasidiomycetes are of special interest in biotechnological research for their versatile potential in the degradation of lignocellulosic biomass, chiefly attributed to ligninolytic enzymes along with exo, endo β-glucanases, xylanases, esterases, pectinases, mannanases, cellobiohydrolases, polysaccharide monooxygenases. Relatively little is known about the metabolic process and the subsequent polysaccharide degradation. Transcriptomic analysis of lignicolous fungi grown on different substrates, although attempted by researchers, has focused on a fairly small group of species reporting the expression of fungal genes in response to lignocellulosic biomass as a substrate. This study accordingly reports analysis of transcriptome of a white-rot Basidiomycete L.squarrosulus grown in simple potato dextrose broth supplemented with aromatic compound, reactive black dye to gain an insight into the degradation ability of the fungus. RNA was sequenced using Illumina NextSeq 500 to obtain 6,679,162 high-quality paired-end reads that were assembled de novo using CLC assembly cells to generate 25,244 contigs. Putative functions were assigned for the 10,494 transcripts based on sequence similarities through BLAST2GO 5.2 and Function annotator.ResultsFunctional assignments revealed enhanced oxidoreductase activity through the expression of diverse biomass-degrading enzymes and their corresponding coregulators. CAZyme analysis through dbCAN and CUPP revealed the presence of 6 families of polysaccharide lyases, 51 families of glycoside hydrolases, 23 families of glycoside transferases, 7 families of carbohydrate esterases and 10 families of auxiliary activities. Genes encoding ligninolytic enzymes and auxiliary activities among the transcript sequences were identified through gene prediction by AUGUSTUS and FGENESH. Biochemical analysis of several biomass-degrading enzymes substantiated the functional predictions.ConclusionIn essence, L. squarrosulus grown in a simple medium devoid of lignocellulosic substrate demonstrated the presence of a repertoire of lignocellulose-degrading enzymes, simplying that a source of lignocellulose is not required for the expression of these biomass-degrading enzymes. This study on the transcriptome analysis of L. squarrosulus revealed significant facts on this front and will definitely enhance the knowledge about the biodegradative ability of this fungus, potentially paving the way for efficient biotechnological applications utilizing its potency in biomass degradation and its future functional exploitation in biomass conversion applications.

Genomic insights into the phylogeny and biomass-degrading enzymes of rumen ciliates

Understanding the biodiversity and genetics of the gut microbiome has important implications for host physiology. One underexplored and elusive group is ciliated protozoa, which play crucial roles in regulating gut microbial interactions. Integrating single-cell sequencing and an assembly-and-identification pipeline, we acquired 52 high-quality ciliate genomes of 22 rumen morphospecies for all major abundant clades. With these genomes, we firstly resolved the taxonomic and phylogenetic framework that reclassified them into 19 species spanning 13 genera and reassigned the genus Dasytricha from Isotrichidae to a new family Dasytrichidae. Via extensive horizontal gene transfer and gene family expansion, rumen ciliates possess a broad array of enzymes to synergistically degrade plant and microbial carbohydrates. In particular, ~80% of the degrading enzymes in Diplodiniinae and Ophryoscolecinae act on plant cell wall, and the high activities of their cellulase, xylanase and lysozyme reflect the potential of ciliate enzymes for biomass-conversion. Additionally, the new ciliate dataset greatly facilitated the rumen metagenomic analyses by allowing ~12% of reads to be classified.

Matrix Effectors and Cancer

Extracellular matrices (ECMs) are highly dynamic three-dimensional structural meshworks composed of macromolecules, such as proteoglycans/glycosaminoglycans (PGs/GAGs), collagens, laminins, elastin, (glyco)proteins, and matrix-degrading enzymes, such as proteases and glycosidases [...]

Directed Evolution of an Efficient and Thermostable PET Depolymerase

The recent discovery of a hydrolytic enzyme, IsPETase, that can deconstruct poly(ethylene) terephthalate (PET), has sparked great interest in biocatalytic approaches to recycle plastics. Realisation of commercial utility will require the development of robust engineered enzymes that meet the demands of industrial processes. Although rationally engineered variants of PETases have been reported, enzymes that have been experimentally optimised through iterative rounds of directed evolution - the go-to method for engineering industrially useful biocatalysts – have not yet been described. Here, we report the development and implementation of an automated, high-throughput directed evolution platform for engineering polymer degrading enzymes. Evaluation of >13,000 IsPETase variants, applying catalytic activity at elevated temperatures as a primary selection pressure, afforded a HotPETase variant with 21 mutations that has a melting temperature of 82.5C and can therefore operate near or above the glass transition temperature of PET (60-70C). HotPETase can depolymerise semi-crystalline PET more rapidly than previously reported PETases and can selectively deconstruct the PET component of a laminated packaging multi-material. Structural characterisation of HotPETase reveals several interesting features that have emerged during evolution to improve thermotolerance and catalytic performance. Our study establishes laboratory evolution as a platform to engineer useful plastic degrading enzymes to underpin biocatalytic plastic recycling processes.

Modulation of sensory perception by hydrogen peroxide enables Caenorhabditis elegans to find a niche that provides both food and protection from hydrogen peroxide

Hydrogen peroxide (H2O2) is the most common chemical threat that organisms face. Here, we show that H2O2 alters the bacterial food preference of Caenorhabditis elegans, enabling the nematodes to find a safe environment with food. H2O2 induces the nematodes to leave food patches of laboratory and microbiome bacteria when those bacterial communities have insufficient H2O2-degrading capacity. The nematode’s behavior is directed by H2O2-sensing neurons that promote escape from H2O2 and by bacteria-sensing neurons that promote attraction to bacteria. However, the input for H2O2-sensing neurons is removed by bacterial H2O2-degrading enzymes and the bacteria-sensing neurons’ perception of bacteria is prevented by H2O2. The resulting cross-attenuation provides a general mechanism that ensures the nematode’s behavior is faithful to the lethal threat of hydrogen peroxide, increasing the nematode’s chances of finding a niche that provides both food and protection from hydrogen peroxide.